Hepsin (also known as TMRPRSS1) is a cell surface expressed chymotrypsin-like serine protease and a member of the family of type II transmembrane serine proteases (TTSP), which also include matriptase (also known as MT-SP1) and enteropeptidase (1). The human hepsin gene, located on chromosome 19 at q11-13.2 (2), encodes a 417 amino acid polypeptide (3) comprised of a short N-terminal cytoplasmic tail, a transmembrane region and an extracellular domain (Arg45-Leu417) composed of the scavenger receptor cysteine-rich (SRCR) and protease domains. Hepsin zymogen is activated autocatalytically by cleavage at Arg162-Ile163 (4), forming a heterodimeric enzyme with the protease domain disulfide-linked (Cys153-Cys277) to the SRCR domain. In addition to the covalent Cys-Cys bond, the recently determined crystal structure of hepsin revealed that SRCR and protease domains share an extensive interface region, each domain burying about 1200 Å2 (5). Because this interface region is located near membrane-proximal residues of the SRCR domain, the hepsin protease domain and active site may be positioned close to the cell surface (5). This is fundamentally different from other cell surface-assembled serine proteases, such as coagulation factors VIIa (FVIIa)1, IXa and Xa, whose active sites are located far above (60-80 Å) the membrane surface (6-8).
The physiological function of hepsin has been elusive. Except for coagulation factor VII, no macromolecular substrates are known and physiologically relevant inhibitors have not been identified. A role of hepsin in blood coagulation was suggested by Kazama et al. (1995) (9) demonstrating that hepsin-transfected cells can activate coagulation factor VII. However, hepsin-deficient mice were viable and showed no blood coagulation disorders (10,11), casting doubt on the importance of hepsin in normal hemostasis. However, it is possible that hepsin may contribute to fibrin formation in pathologic situations, such as renal cell carcinoma (12), where the primary initiator of blood coagulation, tissue factor, (13,14), is absent. Furthermore, other studies have suggested a functional link between hepsin and cellular growth. Depending on the tumor cell line and experimental conditions used, hepsin has been reported to have growth promoting (15) or growth suppressing activity (16). Additional information regarding hepsin can be found in, inter alia, PCT Pub. No. WO2004/009803; U.S. Pat. Nos. 6,482,630; 6,423,543; 5,981,830; U.S. Pat. Appl. Pub. No. 2004/0009911 A1; U.S. Pat. Appl. Pub. No. 2004/0001801 A1; U.S. Pat Appl. Pub. No. 2003/0223973 A1; U.S. Pat. Appl. Pub. No. 2003/0175736 A1; U.S. Pat. Appl. Pub. No. 2003/0013097 A1 (also WO02/059373); U.S. Pat. Appl. Pub. No. 2003/0049645 (also WO02/064839); and U.S. Pat. Appl. Pub. No. 2004/0132156.
Recent gene expression experiments identified hepsin as one of the most highly upregulated genes in prostate cancer (17-22). In-situ staining showed hepsin expression on epithelial cells of the prostate secretory glands (19). The expression of hepsin correlated with the neoplastic transformation (19), being highest in tumors of patients with advanced disease and lowest in benign hyperplasia (18,22). In contrast, one study found that low expression of hepsin protein correlated with high Gleason scores and large tumors (20). It is not clear whether this apparent contradiction is related to the methods used, i.e. immunohistochemistry (20) vs. RNA quantification (18,22). Furthermore, hepsin is also strongly upregulated in ovarian cancer (23) and in renal cell carcinoma, where it is mainly associated with the epithelial cell type (12).
At the epithelial cell surface, hepsin is ideally situated to interact with components of the extracellular matrix and other membrane associated proteins. Chymotrypsin-like serine proteases, including the TTSP matriptase (synonym MT-SP1) (24,25) which is structurally related to hepsin, are known to activate fibrinolytic enzymes, matrix metalloproteases and latent forms of growth factors, such as hepatocyte growth factor (HGF). HGF promotes cell proliferation, migration, angiogenesis, survival and morphogenesis by activating the receptor tyrosine kinase Met (reviewed in (26,27)). In addition to its importance in normal physiology, the HGF/Met pathway has been implicated in invasive tumor growth and tumor metastasis (26). HGF has high similarity to the serine protease plasminogen and is composed of a α-chain containing an N-domain and four Kringle domains and a β-chain with homology to chymotrypsin-like proteases. It is secreted into the extracellular matrix as an inactive single chain precursor (pro-HGF) and requires activation cleavage at Arg494-Val495 to form the biologically competent, disulfide-linked α/β heterodimer (28-31). This step is mediated by pro-HGF converting serine proteases, such as hepatocyte growth factor activator (HGFA) (32), matriptase (33,34), urokinase-type plasminogen activator (u-PA) (35), factor XIIa (36), factor XIa and plasma kallikrein (34). HGFA and matriptase are inhibited by cell surface-expressed Kunitz-type inhibitors, such as the two hepatocyte growth factor activator inhibitor splice variants HAI-1 (37,38) and HAI-1B (34) and by HAI-2 (39). HAI-2 (also known as placental bikunin) (40) also potently inhibits factor XIa and plasma kallikrein (41), whereas HAI-1B has little or no inhibitory activity (34). Therefore, the biological availability of the pro-HGF pool in the extracellular matrix is regulated by the activities of pro-HGF convertases and their inhibitors.
The expression profile of hepsin in cancer tissues as described above, coupled with its potential role in acting as a regulator of other growth factors the dysregulation of which might underlie carcinogenesis suggests that modulation of hepsin's interaction with its substrate could prove to be an efficacious therapeutic approach. In this regard, there is a clear need to identify hepsin's physiological substrate and/or its physiological modulator(s). The invention fulfills this need and provides other benefits.
All references cited herein, including patent applications and publications, are incorporated by reference in their entirety.